Rhabdoid tumors (RTs) are genomically simple pediatric cancers driven by the biallelic inactivation of SMARCB1, leading to SWI/SNF chromatin remodeler complex deficiency. Comprehensive evaluation of ...the immune infiltrates of human and mice RTs, including immunohistochemistry, bulk RNA sequencing and DNA methylation profiling studies showed a high rate of tumors infiltrated by T and myeloid cells. Single-cell RNA (scRNA) and T cell receptor sequencing highlighted the heterogeneity of these cells and revealed therapeutically targetable exhausted effector and clonally expanded tissue resident memory CD8+ T subpopulations, likely representing tumor-specific cells. Checkpoint blockade therapy in an experimental RT model induced the regression of established tumors and durable immune responses. Finally, we show that one mechanism mediating RTs immunogenicity involves SMARCB1-dependent re-expression of endogenous retroviruses and interferon-signaling activation.
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•Genomically simple RTs are infiltrated by T cell and myeloid populations•Clonally expanded T cell phenotypes suggest a tumor-specific response•Checkpoint blockade induces tumor regression and immune memory in vivo•Endogenous retrovirus expression is linked to the immunogenicity of RTs
Leruste et al. find that, despite their low mutation burden, rhabdoid tumors have a high rate of infiltration by T cells and myeloid cells, and the immunogenicity is linked to endogenous retrovirus expression. Immune checkpoint blockade induces tumor regression in a rhabdoid tumor mouse model.
High-throughput RNA-sequencing has become the gold standard method for whole-transcriptome gene expression analysis, and is widely used in numerous applications to study cell and tissue ...transcriptomes. It is also being increasingly used in a number of clinical applications, including expression profiling for diagnostics and alternative transcript detection. However, despite its many advantages, RNA sequencing can be challenging in some situations, for instance in cases of low input amounts or degraded RNA samples. Several protocols have been proposed to overcome these challenges, and many are available as commercial kits. In this study, we systematically test three recent commercial technologies for RNA-seq library preparation (TruSeq, SMARTer and SMARTer Ultra-Low) on human biological reference materials, using standard (1 mg), low (100 ng and 10 ng) and ultra-low (<1 ng) input amounts, and for mRNA and total RNA, stranded and unstranded. The results are analyzed using read quality and alignment metrics, gene detection and differential gene expression metrics. Overall, we show that the TruSeq kit performs well with an input amount of 100 ng, while the SMARTer kit shows decreased performance for inputs of 100 and 10 ng, and the SMARTer Ultra-Low kit performs relatively well for input amounts <1 ng. All the results are discussed in detail, and we provide guidelines for biologists for the selection of an RNA-seq library preparation kit.
Huntington's disease is a fatal neurodegenerative disease characterized by striatal neurodegeneration, aggregation of mutant Huntingtin and the presence of reactive astrocytes. Astrocytes are ...important partners for neurons and engage in a specific reactive response in Huntington's disease that involves morphological, molecular and functional changes. How reactive astrocytes contribute to Huntington's disease is still an open question, especially because their reactive state is poorly reproduced in experimental mouse models. Here, we show that the JAK2-STAT3 pathway, a central cascade controlling astrocyte reactive response, is activated in the putamen of Huntington's disease patients. Selective activation of this cascade in astrocytes through viral gene transfer reduces the number and size of mutant Huntingtin aggregates in neurons and improves neuronal defects in two complementary mouse models of Huntington's disease. It also reduces striatal atrophy and increases glutamate levels, two central clinical outcomes measured by non-invasive magnetic resonance imaging. Moreover, astrocyte-specific transcriptomic analysis shows that activation of the JAK2-STAT3 pathway in astrocytes coordinates a transcriptional program that increases their intrinsic proteolytic capacity, through the lysosomal and ubiquitin-proteasome degradation systems. This pathway also enhances their production and exosomal release of the co-chaperone DNAJB1, which contributes to mutant Huntingtin clearance in neurons. Together, our results show that the JAK2-STAT3 pathway controls a beneficial proteostasis response in reactive astrocytes in Huntington's disease, which involves bi-directional signalling with neurons to reduce mutant Huntingtin aggregation, eventually improving disease outcomes.
We are at a time of considerable growth in transcriptomics studies and subsequent
in silico analysis. RNA sequencing (RNA-Seq) is the most widely used approach to analyse the transcriptome and is ...integrated in many studies.
The processing of transcriptomic data typically requires a noteworthy number of steps, statistical knowledge, and coding skills, which are not accessible to all scientists. Despite the development of a plethora of software applications over the past few years to address this concern, there is still room for improvement.
Here we present DEVEA, an R shiny application tool developed to perform differential expression analysis, data visualization and enrichment pathway analysis mainly from transcriptomics data, but also from simpler gene lists with or without statistical values.
The intuitive and easy-to-manipulate interface facilitates gene expression exploration through numerous interactive figures and tables, and statistical comparisons of expression profile levels between groups. Further meta-analysis such as enrichment analysis is also possible, without the need for prior bioinformatics expertise.
DEVEA performs a comprehensive analysis from multiple and flexible data sources representing distinct analytical steps. Consequently, it produces dynamic graphs and tables, to explore the expression levels and statistical results from differential expression analysis. Moreover, it generates a comprehensive pathway analysis to extend biological insights. Finally, a complete and customizable HTML report can be extracted to enable the scientists to explore results beyond the application. DEVEA is freely accessible at https://shiny.imib.es/devea/ and the source code is available on our GitHub repository https://github.com/MiriamRiquelmeP/DEVEA.
The duplication of genes is one of the main genetic mechanisms that led to the gain in complexity of biological tissue. Although the implication of duplicated gene expression in brain evolution was ...extensively studied through comparisons between organs, their role in the regional specialization of the adult human central nervous system has not yet been well described.
Our work explored intra-organ expression properties of paralogs through multiple territories of the human central nervous system (CNS) using transcriptome data generated by the Genotype-Tissue Expression (GTEx) consortium. Interestingly, we found that paralogs were associated with region-specific expression in CNS, suggesting their involvement in the differentiation of these territories. Beside the influence of gene expression level on region-specificity, we observed the contribution of both duplication age and duplication type to the CNS region-specificity of paralogs. Indeed, we found that small scale duplicated genes (SSDs) and in particular ySSDs (SSDs younger than the 2 rounds of whole genome duplications) were more CNS region-specific than other paralogs. Next, by studying the two paralogs of ySSD pairs, we observed that when they were region-specific, they tend to be specific to the same region more often than for other paralogs, showing the high co-expression of ySSD pairs. The extension of this analysis to families of paralogs showed that the families with co-expressed gene members (i.e. homogeneous families) were enriched in ySSDs. Furthermore, these homogeneous families tended to be region-specific families, where the majority of their gene members were specifically expressed in the same region.
Overall, our study suggests the involvement of ySSDs in the differentiation of human central nervous system territories. Therefore, we show the relevance of exploring region-specific expression of paralogs at the intra-organ level.
Noninvasive biomarkers such as methylated ccfDNA from plasma could help to support the diagnosis of Alzheimer's disease (AD).
A targeted sequencing protocol was developed to identify candidate ...biomarkers of AD in methylated ccfDNA extracted from plasma.
The authors identified differentially methylated CpGs, regions of which were the same as those identified in previous AD studies. Specifically, a differentially methylated CpG of the
gene previously identified in a plasma study of AD was replicated in the study. The
and
regions have been identified in other brain studies of AD and in the authors' study.
Although these biomarkers must be validated in other cohorts, methylated ccfDNA could be a relevant noninvasive biomarker in AD.
We are at a time of considerable growth in the use and development of transcriptomics studies and subsequent
in silico analysis. RNA sequencing is one of the most widely used approaches, now ...integrated in many studies.
The processing of these data may typically require a noteworthy number of steps, statistical knowledge, and coding skills which is not accessible to all scientists. Despite the undeniable development of software applications over the years to address this concern, it is still possible to improve.
Here we present DEVEA, an R shiny application tool developed to perform differential expression analysis, data visualization and enrichment pathway analysis mainly from transcriptomics data, but also from simpler gene lists with or without statistical values.
Its intuitive and easy-to-manipulate interface facilitates gene expression exploration through numerous interactive figures and tables, statistical comparisons of expression profile levels between groups and further meta-analysis such as enrichment analysis, without bioinformatics expertise.
DEVEA performs a thorough analysis from multiple and flexible input data representing distinct analysis stages. From them, it produces dynamic graphs and tables, to explore the expression levels and statistical differential expression analysis results. Moreover, it generates a comprehensive pathway analysis to extend biological insights. Finally, a complete and customizable HTML report can be extracted for further result exploration outside the application. DEVEA is accessible at https://shiny.imib.es/devea/ and the source code is available on our GitHub repository https://github.com/MiriamRiquelmeP/DEVEA.
Abstract
In order to evaluate the immune infiltrate of AT/RT, we conducted a combined immunohistochemical, multiparametric flow cytometry and transcriptomic analysis of a series of 49 human AT/RT. ...This revealed substantial heterogeneity, with some subgroups of AT/RT showing a prominent immune infiltrate. In details, our analyses indicated that: i) myeloid cells were the most abundant immune population, including both microglial cells and non-resident pro-tumoral M2-polarized macrophages, ii) tumor-infiltrating lymphocytes consisted in equal proportions of CD4+ and CD8+ T cells with few regulatory T cells, and iii) immune modulatory molecules PD-L1 and TIM-3 were expressed at high levels. Intratumoral cytolytic activity of CD8+ T cells highly correlated with both interferon gamma and alpha signatures, confirming the involvement of both adaptive and innate immune cells in the anti-tumor response. A genetically engineered mouse model of AT/RT recently established in our laboratory recapitulates many of these traits, including important infiltration by both lymphoid and myeloid cells sharing similar phenotypic characteristics. Using this model, we showed that blockade of the PD-1/PD-L1 pathway significantly impaired tumor growth and induced memory against a second engraftment, confirming an anti-tumoral T cell memory response. Combination with TIM-3 blockade showed synergistic effects. Additionally, targeting the myeloid infiltrate by TLR3 activation with poly(I:C) induced a potent anti-tumor effect which, combined with PD-1 blockade, led to complete tumor regression in over 85% of treated mice. In conclusion, we demonstrate that immune infiltration is a recurrent property of AT/RT and that immunotherapy, particularly combination regimens, have promising therapeutic potential.
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